Alcohol is the most commonly used and abused drug in the United States. Deleterious alcohol-related health effects, attributed to internal organ toxicity, include irreversible brain tissue injury. Brain tissue of chronic alcoholics features neurodegeneration paralleling neuro-cognitive deficits. The causes of HIV-1-associated neurotoxicity, clinically manifesting as HIV-1 associated dementia (HAD), include excitotoxic effects of glutamate, secretory products of chronically activated glial cells, and oxidative stress, similar culprits to ones mediating alcohol-induced neuronal injury. Alcohol abuse and HIV-1 infection of central nervous system (CNS) could result in combined toxic effects leading to neuronal demise and cognitive dysfunction. The current grant application is focused on putative mechanisms of enhanced neurotoxicity in the setting of alcohol abuse and HIV-1 CNS infection. Specifically, we will study unique aspects of astrocyte dysfunction caused by HIV-1 CNS infection and alcohol abuse. We hypothesize that astrocyte dysfunction caused by alcohol metabolites and oxidative stress results in (1) increased glutamate levels via down regulation of excitatory amino acid transporter (EAAT-2, the primary astrocyte glutamate scavenger) causing neuronal injury; (2) production of pro-inflammatory factors (via activation of Src kinases and phospholipase A2); and (3) enhanced activity of metalloproteases (MMPs) resulting in loss of blood brain barrier (BBB) integrity. We mechanistically address the role of oxidative stress in astrocytes leading to production of pro-inflammatory molecules and impairment of glutamate uptake by astrocytes. Using a combination of in vitro systems, an HIVE animal model chronically exposed to alcohol, and in vivo magnetic resonance spectroscopy, we will investigate the astrocyte dysfunction as a focal point of combined effects of HIV-1 and alcohol abuse in the CNS. We will address the following questions: (1) What are the underlying mechanisms causing astrocyte pro-inflammatory phenotype in the setting of alcohol abuse and HIV-1 CNS infection that cause activation of MMPs and BBB dysfunction? (Aim 1); (2) What is the role/contribution of enhanced production of reactive oxygen species, Rho GTPases, and NF-?B signaling in diminished expression and function of EAAT-2 in astrocytes? (Aim 2); (3) How effective are the therapeutics that target oxidative stress and EAAT-2 dysregulation in ameliorating neurotoxicity and BBB impairment in an animal model for HIV-1 encephalitis and alcohol abuse? (Aim 3). Antioxidants and specific signaling inhibitors will be utilized to delineate pathways involved in these effects. We believe that the proposed works are highly significant as they will uncover novel mechanisms involved in the combined effects of HIV-1 and alcohol abuse in the CNS and propose therapeutic approaches based on these investigations. Public Health Relevance: Alcohol is abused by millions of Americans has long lasting toxic effects on the central nervous system. Clinical studies indicated that alcohol dependence has an additive effect on cognitive deficits associated with HIV-1 infection. Current proposal aims to understand mechanisms of combined effects of HIV-1 and alcohol abuse in the brain and to propose neuroprotective therapies.